Cloning and Haplotype Analysis of GSN4, a Gene for Grain Size and Grain Number in Rice

doi:10.16819/j.1001-7216.2026.251212

Abstract

Abstract: 【Objective】Grain size is a critical determinant of rice yield. This study aims to explore novel genes regulating grain size and expand the current genetic regulatory network, and provide valuable genetic resources for molecular design breeding of high-yield rice varieties.【Method】The large-grain mutant gsn4, generated by EMS mutagenesis of the japonica rice variety W7, was used as experimental material. A combination of systematic phenotypic identification, cytological observation, gene mapping, functional verification, subcellular localization, and haplotype analysis was performed to clone and functionally characterize the candidate gene.【Result】 Phenotypic identification revealed that the gsn4 mutant exhibited pleiotropic effects. Compared with the wild type, its grain length, grain width, and 1000-grain weight were significantly increased, whereas plant height, panicle length, and grain number per panicle were significantly decreased. Cytological analysis indicated that GSN4 regulates rice grain size by modulating both cell proliferation and expansion in glumes. Map-based cloning, together with CRISPR/Cas9 gene-editing validation, confirmed LOC_Os04g46450 as the candidate gene underlying GSN4. This gene encodes a nuclear-localized E3 ubiquitin ligase protein, which acts as a negative regulator of rice grain size and modulates plant growth and development through the brassinosteroid (BR) biosynthesis pathway. Haplotype analysis further identified Hap1 as the elite haplotype conferring the most favorable effects on grain width and 1000-grain weight. 【Conclusion】Collectively, this study elucidates the function and regulatory mechanism of GSN4, enriches the genetic regulatory network governing rice grain size, and provides important genetic resources and a theoretical basis for molecular design breeding of high-yield rice varieties.

Key words: grain size, GSN4, cell cycle, cell expansion, rice, haplotype, BR

摘要： 【目的】粒形是决定水稻产量的重要因素之一，挖掘水稻粒形调控新基因，完善水稻粒形遗传调控网络，为高产水稻分子设计育种提供基因资源。【方法】以EMS诱变粳稻品种武运粳7号（W7）获得的大粒突变体gsn4为试验材料，系统开展表型鉴定、细胞学观察、基因定位、功能验证、亚细胞定位及单倍型分析等对候选基因进行克隆和功能分析。【结果】表型鉴定表明，gsn4突变体表现出一因多效性，与野生型相比，其粒长、粒宽及千粒重显著增加，而株高、穗长及每穗粒数显著降低；细胞学分析显示，gsn4通过影响颖壳细胞的增殖与扩张过程调控水稻籽粒大小。图位克隆结合CRISPR/Cas9基因编辑验证，明确LOC_Os04g46450为GSN4的候选基因，该基因编码核定位的E3泛素连接酶蛋白，属于水稻粒形负调控因子，通过参与油菜素内酯(BR)生物合成通路调控水稻生长发育进程。单倍型分析结果筛选出对粒宽和千粒重调控效果最优的优异单倍型Hap1。【结论】本研究明确了GSN4的功能及调控机制，丰富了水稻粒形遗传调控网络，为高产水稻品种的分子设计育种提供了重要的基因资源和理论依据。

关键词: 粒形, GSN4, 细胞周期, 细胞扩张, 水稻, 单倍型, 油菜素内酯

CHAI Bingze, WANG Junge, WU Kaixiong, LIU Jialong, WU Yue, WANG Huisen, TAN Lin, REN Deyong1, GAO Zhenyu, ZHU Li, ZHANG Qiang, ZHANG Guangheng, QIAN Qian, HU Jiang. Cloning and Haplotype Analysis of GSN4, a Gene for Grain Size and Grain Number in Rice[J]. Chinese Journal OF Rice Science, 2026, 40(5): 584-597.

柴炳泽, 王俊格, 吴凯雄, 刘嘉龙, 吴越, 王荟森, 谭霖, 任德勇, 高振宇, 朱丽, 张强, 张光恒, 钱前, 胡江. 水稻粒形和穗粒数基因GSN4的克隆与单倍型分析[J]. 中国水稻科学, 2026, 40(5): 584-597.

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